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Functional expression of gastric H,K-ATPase using the baculovirus expression system
C H Klaassen1, T J Van Uem, M P De Moel
1Department of Biochemistry, University of Nijmegen, The Netherlands.
FEBS Letters
|August 30, 1993
Summary
This study developed a new baculovirus vector to express both alpha and beta subunits of gastric H,K-ATPase. Co-expression of both subunits is essential for the enzyme's ATP-dependent phosphorylation activity.
Area of Science:
- Molecular biology
- Biochemistry
- Virology
Background:
- Gastric H,K-ATPase is crucial for acid secretion.
- Understanding the function of its subunits is important for physiological studies.
Purpose of the Study:
- To develop a novel recombinant baculovirus for co-expressing gastric H,K-ATPase alpha and beta subunits.
- To investigate the functional requirement of both subunits for H,K-ATPase activity.
Main Methods:
- Constructed a recombinant baculovirus by replacing polyhedrin and p10 coding sequences.
- Infected Sf9 insect cells with the recombinant baculovirus.
- Assayed ATP-dependent phosphorylation capacity in purified Sf9 membranes.
Main Results:
- The recombinant baculovirus successfully expressed both alpha- and beta-subunits of gastric H,K-ATPase.
- Sf9 cells infected with the virus showed K(+)- and SCH 28080-sensitive ATP-dependent phosphorylation.
- This enzymatic activity was absent when only one subunit was present.
Conclusions:
- Both alpha and beta subunits are essential for the functional activity of gastric H,K-ATPase.
- The novel baculovirus system provides a tool for studying H,K-ATPase function.